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Comparison of angiopoietin-like protein 3 and 4 reveals structural and mechanistic similarities.

ABSTRACT: Elevated plasma triglycerides are a risk factor for coronary artery disease, which is the leading cause of death worldwide. Lipoprotein lipase (LPL) reduces triglycerides in the blood by hydrolyzing them from triglyceride-rich lipoproteins to release free fatty acids. LPL activity is regulated in a nutritionally responsive manner by macromolecular inhibitors including angiopoietin-like proteins 3 and 4 (ANGPTL3 and ANGPTL4). However, the mechanism by which ANGPTL3 inhibits LPL is unclear, in part due to challenges in obtaining pure protein for study. We used a new purification protocol for the N-terminal domain of ANGPTL3, removing a DNA contaminant, and found DNA-free ANGPTL3 showed enhanced inhibition of LPL. Structural analysis showed that ANGPTL3 formed elongated, flexible trimers and hexamers that did not interconvert. ANGPTL4 formed only elongated flexible trimers. We compared the inhibition of ANGPTL3 and ANGPTL4 using human very-low-density lipoproteins as a substrate and found both were noncompetitive inhibitors. The inhibition constants for the trimeric ANGPTL3 (7.5 ± 0.7 nM) and ANGPTL4 (3.6 ± 1.0 nM) were only 2-fold different. Heparin has previously been reported to interfere with ANGPTL3 binding to LPL, so we questioned if the negatively charged heparin was acting in a similar fashion to the DNA contaminant. We found that ANGPTL3 inhibition is abolished by binding to low-molecular-weight heparin, whereas ANGPTL4 inhibition is not. Our data show new similarities and differences in how ANGPTL3 and ANGPTL4 regulate LPL and opens new avenues of investigating the effect of heparin on LPL inhibition by ANGPTL3.

SUBMITTER: Gunn KH

PROVIDER: S-EPMC7949051 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundThe majority of chicken microbiota studies have used the ceca as a sampling site due to the specific role of ceca in chicken productivity, health and wellbeing. However, sampling from ceca and other gastrointestinal tract sections requires the bird to be sacrificed. In contrast, fecal sampling does not require sacrifice and thus allows the same bird to be sampled repeatedly over time. This is a more meaningful and preferred way of sampling as the same animals can be monitored and tracked for temporal studies. The commonly used practice of selecting a subset of birds at each time-point for sacrifice and sampling introduces added variability due to the known animal to animal variation in microbiota.ResultsCecal samples and fecal samples via cloacal swab were collected from 163 birds across 3 replicate trials. DNA was extracted and 16S rRNA gene sequences amplified and pyrosequenced to determine and compare the phylogenetic profile of the microbiota within each sample. The fecal and cecal samples were investigated to determine to what extent the microbiota found in fecal samples represented the microbiota of the ceca. It was found that 88.55% of all operational taxonomic units (OTUs), containing 99.25% of all sequences, were shared between the two sample types, with OTUs unique for each sample type found to be very rare. There was a positive correlation between cecal and fecal abundance in the shared sequences, however the two communities differed significantly in community structure, represented as either alpha or beta diversity. The microbial populations present within the paired ceca of individual birds were also compared and shown to be similar.ConclusionsFecal sample analysis captures a large percentage of the microbial diversity present in the ceca. However, the qualitative similarities in OTU presence are not a good representation of the proportions of OTUs within the microbiota from each sampling site. The fecal microbiota is qualitatively similar to cecal microbiota but quantitatively different. Fecal samples can be effectively used to detect some shifts and responses of cecal microbiota.

Project description:BackgroundExperimental determination of protein 3D structures is expensive, time consuming and sometimes impossible. A gap between number of protein structures deposited in the World Wide Protein Data Bank and the number of sequenced proteins constantly broadens. Computational modeling is deemed to be one of the ways to deal with the problem. Although protein 3D structure prediction is a difficult task, many tools are available. These tools can model it from a sequence or partial structural information, e.g. contact maps. Consequently, biologists have the ability to generate automatically a putative 3D structure model of any protein. However, the main issue becomes evaluation of the model quality, which is one of the most important challenges of structural biology.ResultsGOBA--Gene Ontology-Based Assessment is a novel Protein Model Quality Assessment Program. It estimates the compatibility between a model-structure and its expected function. GOBA is based on the assumption that a high quality model is expected to be structurally similar to proteins functionally similar to the prediction target. Whereas DALI is used to measure structure similarity, protein functional similarity is quantified using standardized and hierarchical description of proteins provided by Gene Ontology combined with Wang's algorithm for calculating semantic similarity. Two approaches are proposed to express the quality of protein model-structures. One is a single model quality assessment method, the other is its modification, which provides a relative measure of model quality. Exhaustive evaluation is performed on data sets of model-structures submitted to the CASP8 and CASP9 contests.ConclusionsThe validation shows that the method is able to discriminate between good and bad model-structures. The best of tested GOBA scores achieved 0.74 and 0.8 as a mean Pearson correlation to the observed quality of models in our CASP8 and CASP9-based validation sets. GOBA also obtained the best result for two targets of CASP8, and one of CASP9, compared to the contest participants. Consequently, GOBA offers a novel single model quality assessment program that addresses the practical needs of biologists. In conjunction with other Model Quality Assessment Programs (MQAPs), it would prove useful for the evaluation of single protein models.

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Comparison of angiopoietin-like protein 3 and 4 reveals structural and mechanistic similarities.

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